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Methods and Systems for Purifying Non-Complexed Botulinum Neurotoxin

Inactive Publication Date: 2011-04-21
REVANCE THERAPEUTICS INC
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  • Abstract
  • Description
  • Claims
  • Application Information

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Benefits of technology

[0013]In some embodiments, the sample is subjected to a nuclease digestion before loading on the hydrophobic interaction column. Preferably, the nuclease is derive

Problems solved by technology

C. botulinum and its spores commonly occur in soil and putrefying animal carcasses, and can grow in improperly sterilized or improperly sealed food containers, which are the cause of many botulism cases.
Botulism symptoms can include difficulty walking, swallowing, and speaking, and can progress to paralysis of the respiratory muscles and finally death.
Such approaches have provided relatively low yields, however, typically less than about 10%.
These approaches, however, require extra steps to reform a complete and biologically active botulinum toxin protein.

Method used

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  • Methods and Systems for Purifying Non-Complexed Botulinum Neurotoxin

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Comparison of Inventive Process with a Modified Schantz Process

[0077]Purifications of non-complexed botulinum toxin type A using processes within the scope of the instant invention (‘inventive process“) were directly compared to purifications based on the traditional Schantz approach, further modified by the addition of chromatographic steps to provide the non-complexed form (Modified Schantz process”). Briefly, Clostridium botulinum bacteria were cultured and allowed to grow until fermentation was complete (usually about 72 to about 120 hours from inoculation to harvest). A volume of 30 L of the fermentation culture then was used in each of the following purification procedures.

[0078]The modified Schantz process used involved typical acidification of the fermentation culture to precipitate the toxin, followed by ultramicrofiltration (UF) and diafiltration (DF) to concentrate the raw toxin. DNase and RNase were added to the harvested toxin to digest (hydrolyze) nucleic acids, which ...

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Abstract

Methods and systems for chromatographically purifying a botulinum neurotoxin are provided. These methods and systems allow for efficient purification of a non-complexed form of the botulinum neurotoxin in high purity and yield that can be used as an active ingredient in pharmaceutical preparations.

Description

[0001]This application claims the benefit of priority to U.S. Provisional Application No. 61 / 253,810, filed on Oct. 21, 2009. The contents of this U.S. Provisional Application are hereby incorporated by reference in their entirety.FIELD OF THE INVENTION[0002]This invention relates generally to chromatographic methods and systems for purifying free botulinum neurotoxin from cell cultures to produce a high purity, high potency product.BACKGROUND OF THE INVENTION[0003]Botulinum toxin is a neurotoxic protein produced by the bacterium Clostridium botulinum, as well as other Clostridial species, such as Clostridium butyricum, and Clostridium baraffi. The toxin blocks neuromuscular transmission and causes a neuro-paralytic illness in humans and animals, known as botulism. C. botulinum and its spores commonly occur in soil and putrefying animal carcasses, and can grow in improperly sterilized or improperly sealed food containers, which are the cause of many botulism cases. Botulism symptoms...

Claims

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Application Information

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IPC IPC(8): C07K1/18
CPCC12N9/52C07K14/33C12Y304/24069
Inventor RUEGG, CURTIS L.
Owner REVANCE THERAPEUTICS INC
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